Propolis: anti-Staphylococcus aureus activity and synergism with antimicrobial drugs

Propolis is a natural resinous substance collected by bees from tree exudates and secretions. Its antimicrobial activity has been investigated and inhibitory action on Staphylococcus aureus growth was evaluated The in vitro synergism between ethanolic extract of propolis (EEP) and antimicrobial drugs by two susceptibility tests (Kirby and Bauer and E-Test) on 25 S. aureus strains was evaluated Petri dishes with sub-inhibitory concentrations of EEP were incubated with 13 drugs using Kirby and Bauer method and synergism between EEP and five drugs [choramphenicol (CLO), gentamicin (GEN), netilmicin (NET), tetracycline (TET), and vancomycin (VAN)] was observed. Nine drugs were assayed by the E-test method and five of them exhibited a synergism [CLO, GEN, NET, TET, and clindamycin (CLI)]. The results demonstrated the synergism between EEP and antimicrobial drugs, especially those agents that interfere on bacterial protein synthesis.

Morphology and protein patterns of honey bee drone accessory glands.

We used light and transmission electron microscopy to examine the morphology of the accessory glands of immature and mature adult males of Apis mellifera L. We also made an electrophoretic analysis of the protein content of the mature gland. The glands of the immature male actively secrete a mucous substance that can be seen in the lumen of the gland of the mature male. This secretion stains with mercury bromophenol blue and with periodic acid-Schiff reaction, which stain glyconjugates. The protein content was higher in the lumen secretion than in the gland wall extracts. The electrophoresis patterns of the wall extracts were different from those of the secretion found in the gland lumen.

Hydrolases in the hypopharyngeal glands of workers of Scaptotrigona postica and Apis mellifera (Hymenoptera, Apinae)

Hydrolytic enzymes from hypopharyngeal gland extracts of newly emerged, nurse and foraging workers of two eusocial bees, Scaptotrigona postica, a native Brazilian stingless bee, and the Africanized honey bee (Apis mellifera) in Brazil, were compared. The hypopharyngeal gland is rich in enzymes in both species. Fifteen different enzymes were found in the extracts, with only a few quantitative differences between the species. Some of the enzymes present in the extracts may have intracellular functions, while others seem to be digestive enzymes. Scaptotrigona postica, had lower β-glucosidase and higher lipase esterase activities than A. mellifera. The differences may be due to different feeding habits and behavioral peculiarities of the two species. ©FUNPEC-RP.

Africanized honeybees: Biological characteristics, urban nesting behavior and accidents caused in Brazilian cities (Hymenoptera: Apidae)

In 1956 African honeybee queens (Apis mellifera scutellata) were imported from South Africa and Tanzania to Brazil, as part of a government project to increase Brazilian honey production. The European honeybees existing in that country had not adapted well to the tropical conditions and consequently, had a low productivity. The newly introduced bee was known to produce substantially more honey than the other subspecies, but was also famous for its great aggressiveness and quicker attack of intruders with less disturbance. Hoping to create a new hybrid bee that would be both docile and productive, the scientist Warwick Estevam Kerr tried to cross the African and the European subspecies under controlled conditions. However, an accident resulted in the escape of 26 swarms into the Brazilian countryside, where their queens mated with drones of the European resident honeybees. The poly-hybrid bees resulting from these crossings expressed scutellata-like reproductive, foraging, and defensive behaviors and, for this reason, were called Africanized honeybees. They spread rapidly from the introduction area of the African honeybees (near Rio Claro, São Paulo state) to as far south as mid-Argentina and to the north of Texas, also settling in Arizona, New Mexico, California and Nevada, due to their high adaptability to variable ecological conditions. In spite of a few undesirable behaviors, these bees have been invoking larger economic interest because they produce much more honey, have good resistance to diseases and are excellent pollinators. In Brazil, because people frequently disturb the environment, the occupation of urban refuges by Africanized honeybees has been increasing in the last years. The concern with accidents is generally associated with the high swarming frequency recorded during the year and the variety of shelters available in urban areas. This paper deals with the biological characteristics of the Africanized honeybees, their nesting behavior in urban environments, and accidents caused by these bees in Brazilian cities.

Influence of the collection methodology on the Apis mellifera venom composition: Peptide analysis

Differences of venom peptide composition as function of two collection methodologies, electrical stimulation (ES) and reservoir disruption (RD), were analyzed by reverse-phase HPLC in Apis mellifera races - A. m. adansonii, A. m. ligustica and Africanized honeybee. The analyses were performed through determination of the relative number and percentage of each molecular form associated to the peaks eluted by chromatography. Comparison of these profiles revealed qualitative and quantitative differences related to the venom collection methodology as well to the three races analyzed. In contrast to data usually found for venom proteins, the three races presented a major number of peaks or molecular forms when venom was collected by ES. Besides, in general, the relative concentration of each peak was higher for ES in relation to RD. That indicates the presence of molecular precursors in the venom obtained by RD. The presence/absence pattern of the peaks, such as their relative concentrations showed a closer similarity between A. m. adansonii and the Africanized honeybees than that observed between these and A. m. ligustica. The obtained data allowed a discussion about the differences in the relative concentration of each venom component according to the collection methodology, and finally the biological action of the venom in different races. So, these results, apart from being useful to establish a peptide profile for each bee race as a function of the venom collection methodology, pointed out once more that the chromatographic techniques are a great tool for the identification of A. mellifera subspecies.

Composition of stingless bee honey: Setting quality standards

Compositional data from 152 stingless bee (Meliponini) honey samples were compiled from studies since 1964, and evaluated to propose a quality standard for this product. Since stingless bee honey has a different composition than Apis mellifera honey, some physicochemical parameters are presented according to stingless bee species. The entomological origin of the honey was known for 17 species of Meliponini from Brazil, one from Costa Rica, six from Mexico, 27 from Panama, one from Surinam, two from Trinidad & Tobago, and seven from Venezuela, most from the genus Melipona. The results varied as follows: moisture (19.9-41.9g/100g), pH (3.15-4.66), free acidity (5.9-109.0meq/Kg), ash (0.01-1.18g/100g), diastase activity (0.9-23.0DN), electrical conductivity (0.49-8.77mS/cm), HMF (0.4-78.4mg/Kg), invertase activity (19.8-90.1IU), nitrogen (14.34-144.00mg/100g), reducing sugars (58.0-75.7g/100g) and sucrose (1.1-4.8g/100g). Moisture content of stingless bee honey is generally higher than the 20% maximum established for A. mellifera honey. Guidelines for further contributions would help make the physicochemical database of meliponine honey more objective, in order to use such data to set quality standards. Pollen analysis should be directed towards the recognition of unifloral honeys produced by stingless bees, in order to obtain standard products from botanical species. A honey quality control campaign directed to both stingless beekeepers and stingless bee honey hunters is needed, as is harmonization of analytical methods. © 2007 Asociación Interciencia.

Efeito hipocolesterolêmico dos ácidos caféicos da própolis

Natural products have been used in tratamed of hypercholesterolemia. The bee products have been promoting up the man's interest, among them it stands out the propolis, coleted by bees, rich in polyphenols. The biggest polyphenols of propolis was flavonoids and caffeic acids, which have antioxidant power, presenting the protective action to the lipoprotein LDL-cholesterol against the lipid peroxidation. Therefore, the present work was evaluated whether the caffeic acids of the Botucatu's propolis (Botucatu-SP) affect the levels of plasma cholesterol, in rabbits submitted to the rich diet in cholesterol. The animals were divided in three groups: C (n=2) they received commercial ration and water for the whole period; S (n=2) they received normal ration and water in the first period and supplemented ration and water in the second and third periods; S+T (n=5) they received normal ration and water in the first period, supplemented ration and water in the second period and supplemented ration and extract rich in caffeic acids in the third period. The caffeic acids were administered diluted in water, being in the concentration of 0.05g caffeic acid mL -1 kg -1 of animal. Weekly, after fast of 14 hours, the samples of blood were collected from the marginal vein of the ear for determination of the plasma levels of total cholesterol and their fractions. The caffeic acids of the propolis reduced the plasma concentration of total cholesterol in 30% (280 for 199 mg dL -1) in the rabbits treated with flavonoids, while in the animals of the group S those levels were reduced discreetly (380 for 400 mg dL -1). The animals of the group C maintained this biochemical parameter in the normality range during the whole period (50 mg dL -1). Therefore, we concluded that the caffeic acids exert inhibititory activity in the metabolism of the cholesterol, being considered as a substance of action against the hypercholesterolemia.

Response of the stingless bee manduri do mato grosso (Melipona favosa orbignyi, Hymenoptera, Apidae) to variable concentrations of sucrose in artificial feeding sources of different colors

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Éster de sacarose no controle do Varroa estructor em abelhas africanizadas

This study aimed to determine the effect of sucrose ester on the control of Varroa destructor mite infestation in Africanized honeybees. For the in vitro experiments, the product was tested in bees and mites at five concentrations obtained through dilution in water (T0: 100% distilled water; T1: 0.5%; T2: 1%; T3: 2%; T4: 5%; and T5: 10% sucrose ester).For the field studies, the experimental design was completely randomized, with four treatments and seven replicates, totaling 28 colonies, from which seven were the controls, seven were treated with 0.1% sucrose ester, seven with 0.2% sucrose ester, and seven hives with 0.5% sucrose ester diluted in water. In the in vitro study, the sucrose ester at 0.5% concentration caused mite and bee mortality. In the field tests, the product at 0.2% concentration reduced Varroa destructor infestation in Africanized honeybees and, therefore, may be used as a tool to control this pest. At 0.1, 0.2, and 0.5% concentrations, sucrose ester did not impair the establishment of open and capped brood areas, as well as stored food areas in the hive, suggesting it is not toxic to Africanized honeybees.

Propriedade físico-química da própolis em função da sazonalidade e método de produção

The aim of this work was to investigate monthly, for one year the seasonality effect on the physicochemical properties (dry weight, flavonoids contents, pH and antioxidant activity) of ethanolic extract (EAP) of propolis produced by three different techniques (intelligent propolis collector, plastic screen or scraping) fifteen colonies of africanized bees. No differences (p>0.05) in the physicochemical properties due to the technique of propolis production and seasonality were observed.

Chemometric analysis of Hymenoptera toxins and defensins: A model for predicting the biological activity of novel peptides from venoms and hemolymph

When searching for prospective novel peptides, it is difficult to determine the biological activity of a peptide based only on its sequence. The trial and error approach is generally laborious, expensive and time consuming due to the large number of different experimental setups required to cover a reasonable number of biological assays. To simulate a virtual model for Hymenoptera insects, 166 peptides were selected from the venoms and hemolymphs of wasps, bees and ants and applied to a mathematical model of multivariate analysis, with nine different chemometric components: GRAVY, aliphaticity index, number of disulfide bonds, total residues, net charge, pI value, Boman index, percentage of alpha helix, and flexibility prediction. Principal component analysis (PCA) with non-linear iterative projections by alternating least-squares (NIPALS) algorithm was performed, without including any information about the biological activity of the peptides. This analysis permitted the grouping of peptides in a way that strongly correlated to the biological function of the peptides. Six different groupings were observed, which seemed to correspond to the following groups: chemotactic peptides, mastoparans, tachykinins, kinins, antibiotic peptides, and a group of long peptides with one or two disulfide bonds and with biological activities that are not yet clearly defined. The partial overlap between the mastoparans group and the chemotactic peptides, tachykinins, kinins and antibiotic peptides in the PCA score plot may be used to explain the frequent reports in the literature about the multifunctionality of some of these peptides. The mathematical model used in the present investigation can be used to predict the biological activities of novel peptides in this system, and it may also be easily applied to other biological systems. © 2011 Elsevier Inc.

Correlation between mandibular gland secretion and cuticular hydrocarbons in the stingless bee Melipona quadrifasciata

We investigated whether Melipona quadrifasciata worker mandibular gland secretions contribute directly to their cuticular hydrocarbon profile. The mandibular gland secretion composition and cuticular surface compounds of newly emerged worker bees, nurse bees, and foragers were determined by gas chromatography and mass spectrometry and compared. Both the mandibular gland secretions and the cuticular surface compounds of all worker stages were found to be composed almost exclusively of hydrocarbons. Although the relative proportion of hydrocarbons from the cuticular surface and gland secretion was statistically different, there was a high similarity in the qualitative composition between these structures in all groups of bees. © FUNPEC-RP.

Effects of treatment of the fat body trophocytes of Melipona quadrifasciata anthidioides nurse workers and virgin queens in culture by juvenile hormone III and ecdysterone (20-HE)

The fat body (FB) consists of two types of cells: throphocytes and oenocytes. Throphocytes are related to intermediary metabolism storing lipids, carbohydrates, and proteins while oenocytes play role in the lipids and lipoproteins production. The vitellogenin is the precursor of egg yolk (vitelline) and is synthesized on FB. The aim of this work was to analyze the effects of hormones acting in bee reproduction, as juvenile hormone (JH) and ecdisteroids (20 HE) on FB cells, where vitellogenin is synthesized. For the study were chose nurse workers that in Melipona quadrifasciata anthidioides present activated ovaries and produce eggs, and virgin queens whose ovaries are not yet activated, presenting only previtellogenic follicles. FB trophocytes from these classes of bees were cultivated in media containing different amounts of JH and 20-HE. The effects on trophocytes cytoplasm reserves of lipids, proteins, and activity of acid phosphatase were compared by observing preparations from cultured FB, treated and control, by transmission electron microscopy (TEM). The results showed that the hormones effects are related to the bee's caste and functional ovary stage. The role of acid phosphatase on mobilization of the trophocyte reserves was also determined. © 2012 Wiley Periodicals, Inc.

Side-effects of thiamethoxam on the brain andmidgut of the africanized honeybee Apis mellifera (Hymenopptera: Apidae)

The development of agricultural activities coincides with the increased use of pesticides to control pests, which can also be harmful to nontarget insects such as bees. Thus, the goal of this work was assess the toxic effects of thiamethoxam on newly emerged worker bees of Apis mellifera (africanized honeybee-AHB). Initially, we determined that the lethal concentration 50 (LC50) of thiamethoxam was 4.28 ng a.i./μL of diet. To determine the lethal time 50 (LT50), a survival assay was conducted using diets containing sublethal doses of thiamethoxam equal to 1/10 and 1/100 of the LC50. The group of bees exposed to 1/10 of the LC50 had a 41.2% reduction of lifespan. When AHB samples were analyzed by morphological technique we found the presence of condensed cells in the mushroom bodies and optical lobes in exposed honeybees. Through Xylidine Ponceau technique, we found cells which stained more intensely in groups exposed to thiamethoxam. The digestive and regenerative cells of the midgut from exposed bees also showed morphological and histochemical alterations, like cytoplasm vacuolization, increased apocrine secretion and increased cell elimination. Thus, intoxication with a sublethal doses of thiamethoxam can cause impairment in the brain and midgut of AHB and contribute to the honeybee lifespan reduction. © 2013 Wiley Periodicals, Inc.

Cellular responses in the Malpighian tubules of Scaptotrigona postica (Latreille, 1807) exposed to low doses of fipronil and boric acid

Studies of sub-lethal effects of pesticide residues on stingless bees are scarce and morphological analysis of organs would add information to toxicological analysis in order to clarify the continuous exposure of Scaptotrigona postica to insecticides. The aim of this study was to evaluate the morphology and histochemistry of the Malpighian tubules (excretory organ) of S. postica exposed to fipronil or boric acid to detect cellular responses that indicate toxicity or adaptative mechanisms to stress induced by exposure of worker bees to low doses of these chemical compounds. Newly emerged bees were submitted to toxicological bioassays and morphological analyses by optical microscopy and Transmission Electron Microscopy, as well as histochemical methods, were performed to detect proteins and glycoconjugates. Additionally, immunohistochemical detection of DNA fragmentation and HSP70 (70-kDa Heat shock protein) were performed to detect cell death and stress response, respectively. Statistical analysis, for the bioassays conducted with ingestion of contaminated diet with boric acid at 0.75% (w/w) or with fipronil at 0.1μg/kg of food, showed that the survival of bees that ingested the contaminated diets were significantly different to the survival rate presented by the control group (P<0.0001). Although some characteristics indicative of initiation of cell death were observed, the cells remained metabolically active in the processes of excretion and inactivation of chemical compounds. The data from this study reinforce the importance of research on sublethal effects of low doses of pesticides on bees in an attempt to assess a possible realistic dose and evaluate the risk assessment of stingless bee S. postica foraging in the vicinity of cultivated fields and/or in green urban areas. © 2013 Elsevier Ltd.